RNase P is an enzyme present in all living cells. It catalyses the removal of 5′ leader sequences from tRNA precursor molecules. In bacteria, RNase P consists of an RNA molecule of some 400 nt in length (11, 28) and a small (about 120 aa) protein (33). In the division bacteria, the RNA moiety has been shown to function as an efficient catalyst in vitro (12); hence at least in these organisms, RNase P is a ribozyme (an RNA molecule catalysing chemical reactions). Bacterial RNase P RNAs have been separated into two main structural classes. Type A is the most common structural class and type B is found in the low G+C Gram-positive Bacteria (50). The secondary structure of RNase P RNA has been characterised for many bacterial lineages and variation among the helices provides useful phylogenetic information (51).
The RNase P RNA gene sequences are not very well preserved between bacterial groups, (5) but within a genus, genes can be quite similar. Several hundreds of RNase P RNA sequences are present in the RNase P RNA database ((http://jwbrown.mbio.ncsu.edu/RNaseP/home.html).
The order Chlamydiales is a group of obligately intracellular bacteria which have a unique developmental cycle and pathogenicity. They are parasites of humans and a wide variety of animals. Species in the Chlamydiaceae family have recently been reclassified into two genera, Chlamydia and Chlamydophila, that include nine species (43). In addition, new families now also belong to Chlamydiales, and they include Parachlamydiaceae and Simkaniaceae (43). The type species of Parachlamydiaceae is Parachlamydia acanthamoebae, a symbiont of the amoebae Acanthamoeba castellani and an occasional pathogen of people who acquire this amoebae (34). Simkania negevensis is the type species of Simkaniaceae and, like many other chlamydiae, also causes human infection (56, 57, 61).
It has previously been shown that the RNase P RNA genes in the genus Clamydia differ sufficiently between species to be useful as a diagnostic tool (13); thus the gene is potentially useful for strain differentiation. The differences between sequences also give hints to which parts of the molecule are important for catalytic activity, complementing mutational and structural studies.
Another important family of pathogens, where fast and sensitive diagnostic methods are vital, are the mycobacteria. Traditional diagnostic methods have relied on the demonstration of acid-fast bacilli in clinical samples following cultivation. This is reliable but time consuming, since slow-growing species such as Mycobacterium tuberculosis may need six to eight weeks to form a sufficiently large population. In the last few years, many PCR-based detection assays have been developed based on eg the hsp60 gene (16) or the variable interspersing region between the 16S and 23S rRNA genes (15, 24, 30) and this trend continues.
The RNase P RNA gene sequence from M. tuberculosis is known (6) as well as that from M. bovis BCG and M. leprae. The M. bovis sequence is identical to that from M. tuberculosis, while there are differences to M. leprae. The regions in the RNase P RNA gene which have been indicated by other means as important for catalytic activity, were almost totally conserved between the mycobacteria. The close relationship within microbial genuses, such as mycobacteria, has rendered differentiation between species of the same genus very difficult or impossible.